what is an integron? - recinto universitario de...

1

Transferencia Lateral (Horizontal)

de Genes y su Impacto en la

Diversidad Bacteriana

Carlos M. Rodríguez Minguela, Ph.D.

Departamento de Biología

[email protected]

Ext. 3933

What is an integron?

• bacterial gene capture and expression system

• may encode traits that confer selective

advantages

• these may include but are not limited to:

1.antibiotic resistance

3.metabolic genes

2.virulence factors

4.restriction enzymes

2

beta-lactamase

P(int)

P(c)

aminoglycoside

modifying enzymetrimethoprim resistant

dihydrofolate reductase

59-be 59-be 59-be59-be

Structure and function of integrons

integrase

mRNApolycistronic

P(int)

integrase

•excision of captured genes

Function of integrons:

3

P(int)

integrase

•exchange of captured genes among different

integron classes


P(int)

integrase


•reassortment of captured genes

4

beta-lactamase

P(int)

P(c)

aminoglycoside

modifying enzymetrimethoprim resistant

dihydrofolate reductase

59-be 59-be 59-be59-be

Function of integrons

• as reservoirs of cryptic genes

truncated

mRNA

imperfect palindromes (2o structure)

• not expressed

• could be transferred

Function of integrons

• as sources of species genome diversity

E. coli cell A E. coli cell B

Inactivaton of Intl gene:

• results in the fixation of

captured genes

integron

absent

• prevents insertion, rearrangement,

and exicision of genes

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Classification

of

integrons

Two major groups:

1. Mobile integrons (resistance integrons):

• physically linked to mobile DNA elements:

1.transposons 2.conjugative

transposons

3.conjugative

plasmids

• prevalent in clinical strains

• may confer resistance against 9 different antibiotics

6

Two major groups:

2. Superintegrons (chromosomal integrons):

• large numbers of inserted gene cassettes

• not associated with mobile DNA elements

All integrons are grouped into classes

according to the ID% of the integrase

amino acid sequence.

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5’ 3’

5’ 3’

3’

intl3 attl3 blaiIMP-1 attlC ?

5’

Class1

Class 2

Class 3

40%

a.a. ID

61%

a.a. ID

1

Classic resistance (mobile) integrons)

a.a.=amino acid

7

- Integrons are not mobile by themselves.

- they are mobilized by transposons and

conjugative plasmids.

Secuencias de Inserción y Transposones

- elementos genéticos que se mueven

de una molécula de DNA a otra

-pueden moverse de un lugar a otro en un

cromosoma o de un cromosoma a un plásmido

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Secuencias de Inserción y Trasnposones

Secuencias de inserción Transposones

IR

IRIR

IRIRIR

IRIR IR IR

IRIR

•transposón más simple

•tnp: gen de transposasa

reconoce corta y liga DNA durante

transposición

•IR: inverted repeats, punto de

contacto de la transposasa

•se componen de dos secuencias

de inserción

•poseen genes adicionales a la

tnp (transposasa)

Transposición

tnp

tnp

tnp

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Transposons: segments of DNA

capable of jumping from one

DNA molecule into another,

or of moving around to

different positions in the

genome of a single cell.Target DNA Target DNA Target DNA Target DNA IRIR tnp IRIR IRIR tnp IRIRABR gene

Donor DNA IRIR tnp IRIRDonor DNA

Target DNA Target DNA Target DNA ABR gene

Composite transposon

TranspositionTransposition

IS element IS element

Donor DNA IRIR tnp IRIRDonor DNA

Transposition into new target DNA (plasmid, chromosomal)

Transposase

Donor DNA D Donor DNA IRIR tnp IRIR IRIR tnp IRIRABR geneDonor DNA D Donor DNA IRIR tnp IRIR IRIR tnp IRIRABR gene

IS = Insertion Sequence (insertion elements

are the smallest transposons, they

combine to form composite transposons)

IR = inverted repeat, identical

DNA sequences reading in opposite directions

(DNA is double stranded) these are targeted by the transposase

and are compatible with the recipient DNA region

tnp = transposase, transposon

encoded enzyme that excises

the transposon from a donor

DNA molecule and inserts it

into a recipient DNA molecule

Horizontal Gene Transfer

of Antibiotic Resistance

Traits

transposase

integron

integron

integron

tn

tn

tntn

tn

tn

Chromosome

Bacteriophage

Transposon

“Free” DNA

Plasmid

Plasmid

Chromosome

tn

tn

tn

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65% de las diferencias genéticas entre los genomas completos de 70 bacterias

relacionadas se debieron a la presencia de elementos genéticos transmitidos

lateralmente

• genomas de bacteriofagos insertados

• transposones

• secuencias de inserción

Published articles using next generation

high throughput sequencing technology

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Nature 437, 376-380 (15 September 2005)

• novel fibre-optic slide of individual wells

and is able to sequence 25 million bases,

at 99% or better accuracy, in one four-hour run

• 100-fold increase in throughput over

current Sanger sequencing technology

• shotgun sequencing and de novo assembly

of the Mycoplasma genitalium genome with

96% coverage at 99.96% accuracy in one run

of the machine

Beijing Genomics Institute

- reversible terminator-based sequencing chemistry

≥600 Mb/day, using as little as 100 ng to 1ug starting material

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ABI-SOLiD

Output: 6 gigabases

of mappable data.

DNA sequencing

by ligation.

¿Como pueden identificarse

genes que han sido

transferidos horizontalmente?

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• análisis comparativo de genomas

Bioinformática: análisis computacional de secuencias de DNA y proteínas

¿Como pueden identificarse genes que han sido transferidos horizontalmente?

Observaciones que Sugieren Transferencia Lateral de Genes

características atípicas en la secuencia de

un gen en relación con el resto del genoma.

• G+C %

• tendencias en el uso de codones

• presencia de secuencias repetitivas

flanqueando el gen sospechoso


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2. características atípicas en la secuencia de un gen en relación con el resto del

genoma (diferencias en G+C % )


= 65% G+C

= 48% G+C

características atípicas en la secuencia de un gen en relación con el resto del

genoma (tendencias diferentes en el uso de codones )


Pro = CCG

Arg = CGT

= 65% G+C

= 48% G+C

Pro = CCT

Arg = AGA

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